Stem cell reprogramming during oncogenesis and development

项目摘要： 组织干细胞对于补充垂死细胞和伤口修复至关重要。先前 研究已经证明许多癌症可以由干细胞的功能障碍引起， 或者是通过突变的积累，或者是最近的表观遗传变异， 景观Sox 9是一种转录因子，在胚胎发育和发育过程中起重要作用。 维持许多干细胞隔室。在皮肤中，Sox 9是毛囊所必需的 干细胞，而表皮干细胞的特征在于缺乏Sox 9表达。 有趣的是，基底细胞癌（BCC）过表达Sox 9，但已被证明是一种恶性肿瘤。 来源于表皮而不是毛囊干细胞。此外，Sox 9对于BCC是关键的， Sox 9的遗传缺失完全消除了体内肿瘤形成。的机制 异位Sox 9有助于BCC的肿瘤形成，其他癌症仍有待阐明。 然而，在HF中，Sox 9结合并调节关键的开放染色质结构域（超级增强子）。 我的初步研究表明异位表达 Sox 9在成人表皮中的表达通过激活先前沉默的 可能对BCC转化重要的基因。因此，我假设Sox 9基因 通过直接结合和重塑关键染色质在表皮干细胞中的命运开关 增强剂。利用新开发的表达诱导型Sox 9的转基因小鼠， 表皮干细胞，我将绘制和注释染色质和转录变化， 在诱导Sox 9后发生，以确定Sox 9阴性表皮和Sox 9+表皮的偏差 毛囊干细胞利用核小体结合实验的组合， 免疫沉淀和质谱分析，我将确定Sox 9是否可以直接 识别和重塑异染色质，或者它是否通过以下相互作用实现这一点： 其他蛋白质。最后，使用Fuchs实验室开发的一种强大的子宫内慢病毒方法， 将在以下背景下对使用CRISPR/CAS确定的候选人的结果进行功能测试： 肿瘤的形成和发展。我的研究将直接促进我们对Sox 9的了解 介导的染色质重塑和随后的致癌细胞和干细胞活化 转录途径。确定关键的Sox 9靶基因以及相关蛋白 在表皮细胞到基底细胞癌的重编程中， 将肿瘤表观遗传景观“重置”为非恶性形式。

Project Summary: Tissue stem cells are critical for the replenishment of dying cells and for wound-repair. Previous studies have demonstrated that many cancers can arise from the dysfunction of stem cells, either through accumulation of mutations, or more recently, aberrancies in the epigenetic landscape. Sox9, a transcription factor, plays an important role in the development and maintenance of many stem cell compartments. In the skin, Sox9 is essential for hair follicle stem cells, while epidermal stem cells are characterized by lack of Sox9 expression. Interestingly, basal cell carcinomas (BCC) overexpress Sox9, yet have been demonstrated to arise from epidermal and not hair follicle stem cells. Moreover, Sox9 is critical for BCC as genetic loss of Sox9 completely abolishes tumor formation in vivo. The mechanisms by which ectopic Sox9 contributes to tumor formation for BCC and other cancers remain to be elucidated. However, in HFs, Sox9 binds and regulates the key open chromatin domains (super-enhancers) that choreograph stem cell genes, and my preliminary studies indicate that ectopic expression of Sox9 in the adult epidermis shifts the chromatin landscape by activating previously silenced genes that may be important for BCC transformation. Therefore, I hypothesize that Sox9 elicits a fate switch in Epidermal stem cells by directly binding and remodeling chromatin at key enhancers. Utilizing a newly developed transgenic mouse that expresses an inducible Sox9 in the epidermal stem cells, I will map and annotate the chromatin and transcriptional changes that occur after induction of Sox9 to pinpoint deviations from Sox9-negative epidermal and Sox9+ hair follicle stem cells. Exploiting a combination of nucleosome binding experiments, immunoprecipitations and mass spectrometry, I will determine whether Sox9 can directly recognize and remodel heterochromatin, or whether it achieves this through the interactions of other proteins. Finally, using a powerful in utero lentiviral approach developed in the Fuchs lab, I will functionally test the results of candidates identified using CRISPR/CAS in the context of tumor formation and progression. My studies will directly further our knowledge regarding Sox9 mediated chromatin remodeling and subsequent activation of oncogenic and stem cell transcriptional pathways. Identifying the key Sox9 target genes as well as the proteins involved in epidermal to BCC reprogramming will lead to the development of novel therapeutics used to “reset” the tumor epigenetic landscape into a non-malignant form.